As structural biologists tackle ever more challenging systems, the development of efficient methods to deliver large quantities of crystals for X-ray diffraction studies is increasingly important. Proteins that are difficult to crystallize will often produce only small crystals that yield only a few degrees of diffraction data before succumbing to the damaging effects of radiation exposure. For many systems, obtaining a complete dataset to high resolution using very small crystals is possible through the use of microfocus synchrotron beams and the collection and combination of data from multiple crystals. The structural information accessible from very small or very radiation sensitive crystals may be extended through the application of femtosecond crystallography (FX), an emerging method that capitalizes on the extremely bright, short time-scale X-ray pulses produced by X-ray free electron lasers (XFELs). This approach exploits a ‘diffraction before destruction’ phenomenon where a still diffraction pattern is produced by a single X-ray pulse before significant radiation induced electronic and atomic rearrangements occur within the crystal. Since the area of the crystal exposed to the X-ray pulse is completely destroyed after each shot, multiple crystals are required for these experiments. If larger crystals are available, different areas of the crystal may be exposed to obtain multiple stills from a single crystal. In addition, FX confronts another major challenge in structural enzymology by providing a means to determine catalytically accurate structures of radiation sensitive metalloenzymes, which may undergo structural rearrangement upon photo-reduction of the metal center at a synchrotron. In most cases, these experiments also require a large quantity of samples as each area of the crystal can only be exposed once. High throughput crystallization and the implementation of automated sample mounting systems at synchrotron light sources have made data collection from multiple crystals approachable, however challenges still exist. The process of sample exchange using automated mounting systems, which includes mounting a crystal, centering a crystal for data collection, and dismounting the crystal can take between 35 seconds to a few minutes. While this time scale may be suitable for experiments that require screening of at most a few hundred crystals, higher efficiency methods are required for more challenging endeavors at both the synchrotron and XFEL sources. Furthermore, harvesting crystals for data collection can be another time consuming step. Crystal manipulation robots are being developed to automate this process, however crystal harvesting is still primarily done by hand.
One approach for efficient sample delivery and diffraction quality screening is the use of high-density sample containers that hold crystals in known locations coupled with the use of a high-speed sample goniometer for rapid sample positioning. Examples of high-density sample mounting containers for room temperature data collection include microfluidic chips and micro-crystals traps. What is needed is a simple, inexpensive, high density crystal mounting device for data collection at cryogenic or room temperatures, which is compatible with most automated mounting systems and sample storage containers and enables rapid positioning of multiple crystals.